3-desoxy-6alpha-methyl-17alpha-oxygenated-delta4-pregnenes



United States Patent 3,390,157 3-DESOXY-6ot-METHYL-17ot-0XYGENATED-M-PREGNENES Irving Scheer, Somerville, N.J., assignor to OrthoPharmaceutical Corporation, a corporation of New Jersey No Drawing.Filed July 24, 1961, Ser. No. 125,995 7 Claims. (Cl. 260--397.4)

The present invention relates to a new group of esters of6a-methyl-4-pregnen-17a-ol-20-one having the structure in which Y is aradical selected from the group consisting of hydrogen and alkanoylradicals.

The compounds of this invention are valuable pharmaceu-tical agents. Theesters of 6a-methyl-4-pregnen- 17a-ol-20-one possess anti-ovulatoryproperties and are eflective in regulating ovulation. T-he acetate forexample has practically no est-rogenic properties and does not stimulatethe endometrium yet prevents ovulation at lower levels than 17a-ethynylestra-4-en-l7fi-ol-3-one or 170:-ethynyl-S(l0)-estren-l7B-ol-3-one. Moreover the compounds of the presentinvention administered orally prevent ovulation at dosage levels well'below those at which androgenic effects become prominent. The esters ofthe present invention can be administered in conventional dosage formssuch as pills, tablets, capsules, syrups or elixirs for oral use. Theproducts of the present invention may also be compounded in a formsuitable for injection.

It is an object of the present invention to make available an oralcontraceptive that will have minimal side efiects.

The starting material for the compounds of the present invention is6a-methyl-17u-hydroxyprogesterone which is reduced to6x-methyl-4-pregnen-17aol-20'one by the following sequence of reactions:

3,390,157 Patented June 25, 1968 The free alcohol so obtained has littleantiovulatory effect but may be esterified by the procedures describedin the following examples to produce the esters of the presentinvention.

Example I.6u-methyl-4-pregnen-l7a-ol-20.one

A mixture of one gram 6a-methyl-4pregnen-l7u-ol- 3,20-dione, 1 ml. ofethane dithiol, 2 ml. of methylene chloride and 1 gram of pyridinehydrochloride is stirred gently at room temperature for three minutes.The reaction is quenched by adding 30 ml. of methanol and co0ling theflask in an ice bath for 2 minutes. The resulting precipitate isfiltered, washed with ice-cold methanol and dried under vacuum at roomtemperature. The 6ot-methyl- 4-pregnen-l7a-ol20-one-3-thioketal soobtained may be recrystallized from methanol-methylene dichloride togive 0.9 gram of a product melting at 186-188 C. The optical rotation inchloroform 1113 is +99".

Two grams of the 6wmethyl-4-pregnen-17u-ol-20-one- 3-thioketal obtainedas described above is placed in a flask with 200 ml. of ethanol, and 50grams of Raney nickel stored under 95% ethanol is added to the flask.The contents of the flask are refluxed overnight with stirring. Thereaction mixture is then cooled to room temperature and filtered. Thefiltrate is evaporated to dryness under vacuum and the solid residue iscrystallized from methanol-methylene dichloride to yield 0.95 gram of6a-methyl-4-pregnen-l7cx-ol-20 one melting at 190- 193 C. The opticalrotation in chloroform 0: 5 is +51.

Analysis. Calculated for C H ,,O C=79.94, H=1().37. Found: C=79.95, P11031.

Example II.6a-{methyl-4-pregnen-l7u-ol-20-one acetate ExampleIlI.6tx-methyl-4-pregnen-17a-ol-20-0ne propionate To a mixture of 4.0grams 6a-methyl-4-pregnen-l7otol-20-one, 200 ml. propionic acid and 40ml. propionic anhydride is added 4 grams para-toluene sulfonic acid. Aclear solution results within 30 minutes. The solution is permitted tostand overnight at room temperature and is then poured into ice Water.The resulting mixture is extracted with ether and the oil recovered fromthe ether extract is crystallized from methanol. Recrystallization frommethanol-methylene chloride yields 3.8 grams of6a-methyl-4-pregnen-17a-ol-20-one propionate melting at 143-l44. Theoptical rotation in chloroform a is +35 Analysis. Calculated for C H OC=77.67,

TABLE I.COMPARISON OF ANTI-LITTERING EFFECTS IN RATS acetoxyprogesteronotoxyprogesterone, Representative 40 rug/kg. Control 0.5 mg./kg. 1.0mg./kg.

Percent Littering 20 Zero 60 60 Body wt. change, grams +28. 7 +26. +34.0 +25. Significance, p 9 NS 0. 01 NS 1 During first two weeks oftreatment.

2 Not significant.

TABLE II.-COMPARISON 0F ANTI-LITTERING EFFECTS IN MICE3-desoxy-6a-methyl17a-acetoxyprogesterone toxyprogestlfrone,Representative 40 mg./ g. Control mg./kg. 40 nag/kg. 60 mg./kg.

Percent Littering 26. 6 13. 3 Zero 60 66. 7 Body wt. change} grams +0. 8+3. 3 +5.0 +3.0 +2. 1 Significance, p 0. 1 0. 01 0. 001 2 NS 1 Duringfirst two weeks of treatment.

H=9.91. Found: C=77.79, H=9.97.

Example IV.6a-methyl-4-pregnen-l7a-ol-20-one hexanoate Two grams of6tr-methyl-4-pregnen-17a-ol-20one and 42.4 ml. of hexanoic anhyd'rideare placed in a 3-necked flask equipped with a drying tube and nitrogeninlet tube. Two grams paratoluene sulfonic acid is added. After standingone hour at room temperature, a clear solution results.- The solution ispermitted to stand 5 days at room temperature under nitrogen. Tenmilliliters of pyridine is then added and the mixture is steamdistilled. The residue remaining in the distillation flask is extractedwith ether and the ether extract is washed until neutral. The etherextract is dried over sodium sulfate and is then evaporated to dryness.The residual brown oil may be chromatographed on neutral alumina andcrystallized from methanol-methylene dichloride to yield 1.5 grams ofcrystallized 6u-methyl 4 pregnen-l7a-ol-20-one hexanoate melting at78-79. The optical rotation in chloroform D iS +20 Analysis.--Calculatedfor C H O 0:78.45, H: 10.35. Found: C=78.09, H=9.81.

Example V.-6a-methyl-4-pregnen-l7u-ol-20-one cyclopentylpropionate To amixture of 1.0 g. 6a-methyl-4-p-regnen-17a-ol-20- one and 10 ml.cyclopentylpropionic acid is added, drop wise, 4 ml.trifiuoroaceticanhydride. The resulting solution is heated undernitrogen on the steam bath for 15 minutes. The purple solution soobtained is rapidly poured into 500 ml. of a 2% sodium bicarbonatesolution. The mixture is stirred for 30 minutes extracted with methylenedichloride. The methylene dichloride extract is washed until neutralwith 5% aqueous sodium hydroxide then washed with water and dried overanhydrous sodium sulfate. The methylene dichloride solution so obtainedis concentrated and chromatographed on neutral alumina. Thechromatographed solution gave 830 ml. of crystalline6a-methyl-4-pregnen-17ot-ol-20-one cyclopentylpropionate melting at93-95 Analysis-Calculated for G l-1 0 C=79.24, H: 10.20. Found: C=79.01,H=10.27.

An anti-littering assay may be designed to compare the anti-litteringeffect of 3-desoxy-6a-rnethyl-17a-acetoxy progesterone, prepared asdescribed in Example II above, and 3-desoxy-17a-acetoxy-progesterone, byadministering these compounds to adult rats and mice. The sequence ofthe experiment is as follows:

(1) The drug to be evaluated is administered in the die for 7 days withthe sexes segregated.

2 Not significant.

It will be observed from Table I above that the acetate described inExample II reduces littering at doses of 0.5 and 1.0 mg. per kilogram ofbody weight in the rat. The related compound lacking a methyl group inthe 6-position did not impair the fertility of the rat.

From Table II, it will be observed that dosage levels of about 60 mg.per kg. in the mouse are required to prevent littering but the number ofmice producing litters may be reduced by treatment of doses as low as 10mg. per kg. At 40 mg. per kg. the related compound lacking a methylgroup in the 6-position has no apparent effect on the littering of mice.The compound of the present invention is tolerated and allows normalbody weight gain.

The estrogenic properties of the compounds of the present invention asmentioned above are minimal. The estrogenic potency of6a-methyl-4-pregnen47a-ol-20 one acetate may be determined by thestandard assay procedure described by J. S. Evans, R. S. Varney and F.D. Koch in Endocrinology, 28, p. 247 (1941) using the uterine Weightincrease and precocious vaginal opening of immature mice as indications.From the data in Table [III below one may compare the estrogenicity ofthe 3-desoxy- 6a-methyl-l7a-hydroxy progesterone acetate with 3-desoxy-l7a-acetoxy progesterone and 6ot-methyl l7a-hydroxy progesteroneacetate.

TABLE IIL-ORAL ESTROGEN ASSAY Total Number of Ut. Wt. Percent VaginalDosage Animals 0v. Wt. Opening Chi 3-desoxy-l7a-acetoxyprogesteroneControl 5 8. 2:l:1. 3 2. 9:l:0. 4 O 0.1 mg 10 6. 7+0. 47 3. 2&0. 32 01.0 mg 10 6. 5:l:0. 52 2. 7:l:0. 2 0 5.0 mg" 10 6.3i0.4 2. 7:l:0. 17 010.0 mg. 10 7. 3i0. 6 3. 2:l:0. 17 0 3-des0xy-Ga-methyl-li'a-hydroxyprogesterone acetate Control 4 6. 4:1:0. 55 3. 5&0. 28 0 1 mg 9 7.5:h0.4 3.310. 28 0 N.S. 10 mg. 10 8. 6:i:0. 41 3. iii). 17 0 0. 01

fia-methyl-17a-hydroxyprogesterone acetate Control 5 4.610.514 2. 0:l:0.42 0 0.1 mg 1O 5. 6:l:0. 39 2. 5:i:0. 17 0 N.S. 1 0 mg 10 0. 610. 35 2.83:0. 22 0 0. 01 5 0 mg 9 6.810. 49 2. 5:l:0. 31 0 0.01 10. 0 mg 8 8.6:1:0. 5 2. 7:i:0. 22 0 0. 01

3-desoxy-6a-niethyl- Hot-progesterone acetate induces a slight,statistically not significant increase with 1.0 mg.

total dose, but mg. total dose causes an increase of uterine weights,yet it does not affect the vaginal area.

3-desoxy 17a-acetoxyprogesterone fails to stimulate an increase inuterine weights at any of the dose levels tested, (0.1 mg. to 10.0 mg.total dose), and there was no effect on precocious vaginal opening.Ovarian weights remained unchanged.

=6a-methyld7a-hydroxyprogesterone acetate is five to ten times moreuterotrophic than 3-desoxy-6a-methyl- Nix-progesterone acetate since 1.0mg. total dose causes a significant uterine weight increase, yet theresponse curve rises very slowly with increasing doses. Vaginal openingis not affected.

'In general it appears that none of the compounds in Table III aregenuinely estrogenic in view of their low effects on uterine weight andfailure to induce precocious vaginal opening. This response is moretypical for that produced by weak progestational and androgenic agents.More potent androgens such as methyl testosterone with 1.0 mg. dosesproduces a uterine weight of 14 mg. and induces uniform vaginal openingin immature mice.

The androgenicity of the compounds of the present invention may bedetermined by a modification of the technique described by L. G.Henchberger, E. G. Shipley, and R. K. Meyer, Proc. Soc. Exp. Biol. andMed, 83, 170-175 (1953). Immature male rats are castrated and thecompounds are administered by stomach tube starting the following day in7 daily doses. Autopsies are performed on the day after the lasttreatment. The results are shown in Table IV.

TABLE IV.-ORAL ANDROGEN ASSAY3-desoxy-6a-methy1-17a-hydroxy-progesterone-acetate Control..- 10 4.0(6. 5) 7.2 (11.7) 11.4 (18.5) 10.6 (17.0) 0.1 mg 10 3.8 (6.7) 7.0 (12.2) 12.4 (21.8) 8.5 (14.2) 1.0 mg. 10 4.3 (6.9) 6.9 (10.9) 12.0 (19.3)10.9 (17.4) 10.0 mg.-. 9 5.3 (9.8) 9.3 (17.4) 8.3 (15.4) 12.9 (22. 4)20.0 mg... 10 6.6 (10. 9) 13.0 (21.6) 6.0 (9.8) 13.5 (22.0)

6a-methyl-17a-hydroxy-progestcrone acetate Control... 10 3.6 (7.6) 6.2(13.2) 10.7 (22.5) 6.9 (14.2) 0.1 mg.-.- 10 3.9 (8.1) 6.1 (12.8) 10.7(22.0) 7.1 (14. 5) 1.0n1g.--- 9 4.6 (9.1) 7.2 (14.5) 10.8 (21.7) 7.2(14.4) 10.0 mg--- 9 5.1 (9.0) 12.0 (21.3) 7.7 (13. 6) 10.6 (18.6) 20.0mg..- 9 4.7 (8.8) 11.1 (20.9) 6.7 (12.6) 10.6 (19.6)

*Values in parentheses expressed as mg. per 100 g. of body weight.

6a-rnethyl-4-pregnen-17a-ol-20-one on the basis of relative organweights at dose levels up to 1.0 mg. total dose shows no androgenic andanabolic potency and does not affect the adrenals. At 10.0 and 20.0 mg.total dose it exerts an androgenic effect principally on the ventralprostate, but to a lesser degreeon the semi vesicles. The anaboliceifect is only slight.

In comparison 10.0 mg. total dose of methyl testosterone produces arelative weight for ventral prostate of 80 and of the seminal vesiclesof 42 (versus 17 and 10 respectively for 6a-methyl 4 pregnen 17a ol 20oneacetate).

The estrogenic and androgenic effects of desoxy 6arnethyl-lh-hydroxyprogesterone may be compared with the acetic acid ester of this compoundby referring to Tables V and VI.

TABLE V [Estrogeuicity of 3-desoxy-17a-hydroxy progesterone,3-desoxy-6amethyl-17a-hydroxy progesterone acetate and3-Desoxy-6a-mcthyl-17ahydroxy progesterone by subcutaneousadministration 01' 1.5 mg. total dose. (Absolute weights in mg.)]

Ut. Weight Ovarian Wt:

Control 4. 7:1:0. 4 2. 510. 39 3-Desexy-17a-hydr0xy progesterone 5.8:1:0. 32 3. 2:1:0. 26 Control 5. 44:0. 41 2. 8:1:0. 43-Desoxy-6a-methyl-17a-hydroxy progesterone acetate 6. 6:1:0. 47 3. 2&0.3 Control 4. 7:1:0. 4 2. 5i0. 39 3-Desoxy-6a-mcthy1 ydroxy p ogesterone5. 0:1:0). 33 3. 2i0. 22

TABLE VI [Androgenieity of 3desoxy-17a-hydroxy progesterone,3-desoxy-6amethyl-lm-hydroxy progesterone acetate and3-desoxy-6a-rnethyl-17ahydroxy progesterone by subcutaneousadministration of 1.5 mg. total dose of 3 desoxy-17a-hydroxyprogesterone and 3-desoxy-6a-n1ethyl-17ahydroxy progesterone, 5 mg. of3-desoxy-6a-methyl-17a-hydroxy progesterone acetate (Absolute weights inmg.)]

In the literature pertaining to inhibition of ovulation the term"ovulation inhibition is frequently applied to a Wide time-span andphysiological actions that may include inhibition of folliculardevelopment through prolonged administration of inhibiting agents thatgenerally act through inhibition of total ovarian activity, such as thenatural hormones, estrogens, androgens, progestins.

The compounds of the present invention seem to elfect an immediateovulation stop, upon the short term or single administration of theseesters prior to expected ovulation.

The effect described above may be confirmed as follows: Rats are firstchecked for the regularity of their cycles by daily vaginalexaminations. For the test, rats with synchronous cycles are selectedand the drug is administered at the 2 days preceding the day of expectedovulation. The drug is administered either in the diet or by stomachtubing. In the afternoon of the day when ovulation would occur the ratsare autopsied and their fallopian tubes are excised and flushed out torecover ovulated ova which in untreated rats would be located at thispart of the reproductive tract on their migration towards the uterus.Absence of ova is taken as indication that ovulation has been prevented.This is further checked by histological serial sections of the ovaries.If ovulation has been prevented the mature follicles will still containova, and no fresh corpora lutea are present.

In preliminary tests it has been investigated whether or not suchtreatment would merely shift ovulation time either by hastening orpostponing it. Such an effect has not been demonstrated with this typeof compound or schedule.

Compounds that exhibit an anti-ovulatory effect are titrated down to theminimal effective dose exerting this response. At a dosage level of 20=mg./kg. 3-desoxy-17ahydroxy progesterone given in the diet is notcapable of of stopping ovulation since 5 out of 7 rats: ovulated. Whengiven by stomach tube to 10 rats, all but one had ovulated. This resultis equal to that obtained with control animals. Thus 3-desoxy17a-hydroxy progesterone has no direct effect on the process ofovulation.

In contrast to this when 3-desoXy-6a-methyl-17a-hydroxy progesteroneacetate at a dose level of only 1 ung/ kg. is administered by stomachtube to 13 rats, non ovulate. When administered at the same dose level,but only once, on the day preceding proestrus, ovulation is prevented in10 out of 12 rats. When the same amount was administered in the diet,ovulation is prevented in only 7 out of 22 rats.

It is to be understood that the invention is not to be limited to theexact details of operation or exact compounds shown and described asobvious modifications and equivalents will be apparent to one skilled inthe art, and the invention is therefore to be limited only by the scopeof the appended claims.

What is claimed is:

1. A compound having the structure a? CU in which Y is a radicalselected from the group consisting of hydrogen and alkanoyl radicals.

2. A compound having the structure Al '---oooc1n 1 Q 3. A compoundhaving the structure m CU 4. A compound having the structure en. 5. Acompound having the structure (EH;

---on /\l l 25 JHa 6. A compound having the structure ll-n 0:0 l---ooocmornl on; 7. A compound of the formula CH: wherein R is selectedfrom the group consisting of hydrogen and lower alkanoyl.

References Cited UNITED STATES PATENTS 2,715,640 8/1955 Ralls260--397.45 3,033,752 5/1962 Clinton et a1. 167--74 ELBERT L. ROBERTS,Primary Examiner.

M. LIEBMAN, Examiner.

G. E. LANDE, Assistant Examiner.

UNITED STATES PATENT OFFICE. CERTIFICATE OF CORRECTION Patent No.3,390,157 June 25, 1968 Irving Scheer It is certified that error appearsin the above identified patent and that said Letters Patent are herebycorrected as shown below:

Column 4, TABLE III in the heading to the third column thereof, cancel"Percent"; in the heading to the fourth column thereof, "Vaginal" shouldread Percent Vaginal Column 5, TABLE IV, fourth columnyline 3 thereof,"8.6(12.0)" should read 8 .6(13.0) Column 6, line 65 cancel "of";

line 73, "non" should read none Signed and sealed this 20th day ofJanuary 1970.

(SEAL) Attest:

WILLIAM E. SCHUYLER, JR.

Commissioner of Patents Edward M. Fletcher, Jr.

Attesting Officer

1. A COMPOUND HAVING THE STRUCTURE